This is a revised (score 159/26.8%) competing renewal for RO1 CA74730 "Regulation of gammaherpesvirus latency and tumorigenesis" which funded studies of the mechanisms of y-herpesvirus latency using infection with murine gamma-herpesvirus 68 (gammaHV68) as a model. We defined both host immune mechanisms (B cells, antibody, CD8 T cells, IFNalphabeta, STAT1, IFNgamma, perforin, caspase 3, granzymes) and viral genes (M1, M2, gene 50, gene 73 (KSHV LANA homolog), v-bcI-2, v-cyc/in, and v-GPCR) that control persistent replication and latency. Particularly interesting was the role for the v-Bcl-2, which is conserved across gamma-herpesviruses, in latency and persistent infection. We also developed, as proposed, a robust model for gammaHV68 induction of B cell lymphomas. Here we propose to continue identification of the genes critical for latency using newly developed methods for unbiased identification of candidate herpesvirus latency transcripts combined with loss of function mutagenesis. Taking advantage of the new tumor model we will address the fundamentally important question of whether latency and lymphomagenesis require the same genes. To understand the cellular basis of gammaHV68 latency, we will define the effects of gammaHV68 infection and expression of latency associated genes on the proliferation and survival of primary B cells. To understand latency at the molecular level, we will define the mechanisms by which the v-Bcl-2 regulates ?HV68 latency. To this end we have generated new data identifying a host apoptotic pathway (perforin/granzymes) that regulates gammaHV68 latency, performed structural and biochemical characterizations of the KSHV, EBV, and gammaHV68 v-Bcl-2 proteins, and shown that the conserved v-Bcl-2 BH1 domain is important for latency. These data provide a firm foundation for experiments identifying how the v-Bcl-2 protein contributes to chronic ?HV68 infection. These interrelated goals will be pursued through three Aims: Aim 1: Identify the role of latency and tumor associated genes in latency and lymphomagenesis. Aim 2: Identify the role of latency and tumor-associated genes in B cell proliferation and survival. Aim 3: Identify the mechanism by which v-Bcl-2 regulates gammaHV68 latency.